Ion Channel Characterization using Current Voltage Resonance Spectroscopy

使用电流电压共振光谱法表征离子通道

• 批准号: 7915304
• 负责人: Steven Poelzing
• 金额: $ 18.81万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-01 至 2012-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7915304
• 关键词: Action Potentials; Address; Affect; Attenuated; Behavior; Cardiac; Cardiac Myocytes; Cause of Death; Cavia; Cells; Characteristics; Coupled; Data; Diffuse; Disease; Environment; Exhibits; Family; Feedback; Frequencies; Genetic; Heart; Heart failure; Heterogeneity; Individual; Ion Channel; Ions; Kinetics; Left; Measures; Membrane; Membrane Potentials; Methods; Microelectrodes; Modeling; Molecular; Morphology; Muscle Cells; Mutation; Noise; One-Step dentin bonding system; Physiological; Potassium Channel; Predisposition; Proteins; Protocols documentation; Recovery; Relative (related person); Research Personnel; Rest; Right ventricular structure; Signal Transduction; Sodium; Sodium Channel; Spectrum Analysis; Structure; System; Techniques; Testing; Time; Validation; Ventricular; base; density; electric field; electric impedance; indium arsenide; inward rectifier potassium channel; novel; patch clamp; public health relevance; response; simulation; sudden cardiac death; tool; voltage; voltage clamp

DESCRIPTION (provided by applicant): Ion channels of excitable membranes are responsible for synchronizing the firing and recovery of excitable cells such as cardiac myocytes. It is well established that heterogeneities and loss of ion channel function is a major component of lethal diseases such as sudden cardiac death in heart failure and familial forms of genetic ion channel mutations. The behavior of individual ion channels in relatively isolated conditions is well defined due to techniques such as patch clamping which can measure the function of a single ion channel, an ion channel over- expressed in a heterologous system, or an ion channel in a cardiac myocyte under conditions where all other ion channels are suppressed. However, little is known about how ion channels behave as a family under physiological conditions such as a cardiac action potential when more than one ion channel is actively passing current. Specifically, the passage of current by multiple ion channels defines the voltage morphology and contributes to feedback mechanisms activating or in/deactivating other ion channels. The purpose of this proposal is to validate impedance spectroscopy for simultaneously quantifying transsarcolemmal currents INa and IK1 in specific. We chose these two channels based on intriguing previous results our group obtained. In short, the faster conducting right ventricle expresses significantly less Nav1.5 relative to left. We demonstrated that IK1 modulates normal cardiac conduction to a greater extent than Nav1.5. Impedance spectroscopy will be used to demonstrate the feasibility of simultaneously quantifying INa and IK1. In order to address the general hypothesis that each ion channel has a unique characteristic frequency response due to structural differences, the following specific aims will be tested. 1. Determine the characteristic resonant frequency signatures of INa and IK1 in heterologous cells 2. Determine the mechanisms underlying at least one characteristic resonant frequency in sodium and potassium channels 3. Demonstrate that IK1 and INa can be measured simultaneously. In the preliminary data, we now demonstrate that INa and IK1 exhibit similar and unique frequencies that correlate to their respective current amplitudes. Additionally, the preliminary data demonstrates that the time course of the current (INa or IK1) predominates the characteristic frequency response. Therefore, in order to quantify INa or IK1 simultaneously, the predominant signal must be removed by a "difference frequency response correction." Impedance spectroscopy is not new. However, the application of impedance spectroscopy corrected for the predominating signal is a novel method to simultaneously quantify transsarcolemmal ion channels. This is an important tool, because it will allow researchers to finally quantify currents in their native environment when the channels are being affected by the voltage produced by concurrently active channels. Successful completion of this proposal would allow simultaneous quantification of sarcolemmal currents in any excitable cell, not just cardiomyocytes. PUBLIC HEALTH RELEVANCE: Ion channels describe the voltage profile of excitable cardiac cells. When ion channel function changes or becomes heterogeneous between regions of the heart, an individual's susceptibility to sudden cardiac death, the leading cause of death in the U.S., increases significantly. This proposal seeks to develop a method to simultaneously quantify multiple functioning ion channels in cardiac myocytes during a physiologic action potential in order to determine ion channel functional heterogeneity.

描述（由申请人提供）：可兴奋的膜的离子通道负责同步可激发细胞（例如心肌细胞）的发射和恢复。众所周知，异质性和离子通道功能的丧失是致命疾病的主要组成部分，例如心力衰竭中猝死和遗传离子通道突变的家族形式。由于可以在所有其他离子通道抑制所有其他离子通道的条件下，因此可以测量单个离子通道的功能，在异源系统中过度表达的离子通道，在异源系统中过度表达的离子通道的功能，在相对隔离的条件下，单个离子通道在相对孤立的条件下的行为得到了很好的定义。但是，对于在多个离子通道积极传递电流时，在生理条件下（例如心脏动作电位）在生理状况（例如心脏动作电位）中如何表现为家庭。具体而言，多个离子通道的电流通过定义了电压形态，并有助于激活或停用其他离子通道的反馈机制。该提案的目的是验证特定于特定于特异性的跨性膜电流INA和IK1的识别光谱法。我们根据我们的小组获得的先前结果选择了这两个渠道。简而言之，相对于左侧的右心室的速度更快的NAV1.5表达明显较小的NAV1.5。 我们证明，IK1比NAV1.5更大程度地调节正常的心脏传导。 阻抗光谱将用于证明同时量化INA和IK1的可行性。为了解决每个离子通道由于结构差异而具有独特的特征频率响应的总体假设，将测试以下特定目标。 1。确定异源细胞中INA和IK1的特征谐振频率特征。确定在钠和钾通道中至少一种特征共振频率的机制3。证明可以同时测量IK1和INA。在初步数据中，我们现在证明INA和IK1表现出与它们各自的电流振幅相关的相似且独特的频率。此外，初步数据表明，电流（INA或IK1）的时间过程主要是特征频率响应。因此，为了同时量化INA或IK1，必须通过“差异频率响应校正”来删除主要信号。阻抗光谱并不是什么新鲜事物。然而，对主要信号校正的阻抗光谱的应用是一种同时量化跨性别膜离子通道的新方法。这是一个重要的工具，因为它将允许研究人员最终量化其本地环境中的电流，当时通道受到同时有效通道产生的电压的影响。该提案的成功完成将允许在任何令人兴奋的细胞中同时量化肌膜电流，而不仅仅是心肌细胞。公共卫生相关性：离子通道描述了可激发心脏细胞的电压谱。当离子通道功能变化或在心脏区域之间变化或变得异质时，个人对心脏死亡的敏感性（美国的主要死亡原因）显着增加。该建议旨在开发一种在生理动作潜力期间同时量化心肌细胞中多重功能离子通道的方法，以确定离子通道功能异质性。